Lead, ferrochelatase inhibition

Lead-induced anemia results from impairment of heme biosynthesis and acceleration of red blood cell destmction (10,13). Lead-induced inhibition of heme biosynthesis is caused by inhibition of S-aminolevulinic acid dehydratase and ferrochelatase which starts to occur at blood lead levels of 10 to 20 pu gjdL and 25 to 30 //g/dL, respectively (10,13). Anemia, however, is not manifested until higher levels are reached. 

In summary, lead inhibits the activity of certain enzymes involved in heme biosynthesis, namely, 5-aminolevulinic acid dehydratase (ALAD), and ferrochelatase. As a consequence of these changes, heme biosynthesis is decreased and the activity of the rate limiting enzyme of the pathway,... 

Answer C. Lead inhibits both ferrochelatase (increasing the zinc protoporphyrin) and ALA dehydrase (increasing 5-ALA). 

Lead ions also inhibit ferrochelatase-cataiyzed insertion ofFe into protoporphyrin iX in the final step of heme synthesis. 

Formation of heme Uroporphyrinogen III is converted to heme by a series of decarboxylations and oxidations summarized in Figure 21.4. The introduction of Fe2+into protoporphyrin IX occurs spontaneously, but the rate is enhanced by the enzyme ferrochelatase—an enzyme that is inhibited by lead (see p. 279). 

Ferrochelatase and ALA dehydrase are particularly sensitive to inhibition by lead... 

Committed step in heme synthesis, its coenzyme, and inhibitor The committed step in heme synthesis is the formation of 5-amlnolevulinic acid (ALA). The reaction, which requires pyridoxal phosphate as a coenzyme, is catalyzed by ALA synthase. The reaction is inhibited by hemin (the oxidized form of heme that accumulates in the cell when it is being under-used). The conversion of protoporphyrin IX to heme, catalyzed by ferrochelatase, is inhibited by lead. 

Ferrochelatase (protoheme ferro-lyase)401 403 inserts Fe2+ into protoporphyrin IX to form heme. The enzyme is found firmly bound to the inner membrane of mitochondria of animal cells, chloroplasts of plants, and chromatophores of bacteria. While Fe2+ is apparently the only metallic ion ordinarily inserted into a porphyrin, the Zn2+ protoporphyrin chelate accumulates in substantial amounts in yeast, and Cu2+-heme complexes are known (p. 843). Ferrochelatase, whose activity is stimulated by Ca2+, appears to be inhibited by lead ions, a fact that may account for some of the acute toxicity of lead.404... 

A second major lead-induced toxicity involves interruption of heme synthesis. Lead interacts at several steps in the heme biosynthetic pathway (Figure 21.13). As mentioned above, Pb inhibits the enzyme 8-aminolevulinic acid dehydratase (ALA-D), which catalyzes the second step of heme synthesis involving the condensation of two molecules of aminolevulinic acid (ALA) to form porphobilinogen. The result of this inhibition is the accumulation of aminolevulinic acid in the serum and increased excretion of ALA in the urine. A second major disruption of the heme biosynthetic pathway is Pb inhibition of ferrochelatase. This enzyme is responsible for the incorporation of the ferrous ion (Fe2+) into protoporphrin IX to produce heme (Figure 21.2). Accumulated protoporphrin is incorporated into red blood cells and chelates zinc as the cells circulate. This zinc-protoporphrin complex is fluorescent and used to diagnose Pb poisoning. 

Finally, it is widely known that Pb impairs the formation of red blood cells. The mechanism involved in the impairment is that Pb inhibits both 5-aminolevulinic acid dehydratase (ALA-D) (Hernberg et al. 1970) and ferrochelatase (Tephly et al. 1978). These are two key enzymes involved in heme biosynthesis. ALA-D catalyzes the conversion of 5-aminolevulinic acid into porphobilinogen (PBG), whereas ferrochelatase is responsible for catalyzing the incorporation of Fe2+ into protoporphyrin IX to form heme (Figure 9.1). Lead inhibition of the two enzymes appears to be due to its interaction with Zn and Fe required in the process. 

Glyceraldehyde-3-phosphate dehydrogenase, an enzyme in the glycolytic pathway (Chapter 8), is inactivated by alkylation with iodoacetate. Enzymes that use sulfhydryl groups to form covalent bonds with metal cofactors are often irreversibly inhibited by heavy metals (e.g., mercury and lead). The anemia in lead poisoning is caused in part because of lead binding to a sulfhydryl group of fer-rochelatase. Ferrochelatase catalyzes the insertion of Fe2+ into heme. 

Lead interferes with heme pro-duction by inhibiting S-aminole- vulinic acid dehydratase and ferrochelatase. 

Lead has an inhibitory effect on steps in the chain of reactions that lead to the formation of heme, affecting for example the enzymes ALA dehydratase (ALAD) and ferrochelatase (heme synthetase). Lead also inhibits the activity of the enzyme pyrimidine-5-nucleo-tidase (P5N) in red cells. 

Several other classes of proteins have also been implicated as possible targets for lead, including other proteins in the heme biosynthetic pathway, leadbinding proteins in the kidney and brain, and heat shock proteins (342, 500-502). Lead is known to affect several steps in the heme biosynthetic pathway other than that catalyzed by ALAD Other profound effects include stimulation of 5-aminolevulinic acid synthase (ALAS) and decreased levels of iron incorporation into protoporphyrin by ferrochelatase (see Section VI.E.2 and Fig. 34) (10, 503-505). However, not all of these effects are due to direct interactions between lead and enzymes in the heme biosynthetic pathway. For instance, the widespread assertion that lead inhibits ferrochelatase is not supported by studies on the isolated enzyme (506, 507). Furthermore, increased levels of both erythrocyte protoporphyrin IX (EP) and zinc protoporphyrin (ZPP) are observed at high BLLs, suggesting that ferrochelatase is stiU competent to insert zinc into EP and that the increased levels of EP and ZPP associated with lead poisoning are most likely caused by lead interfering with iron uptake or transport (see Sections VI.C.4 and VI.E) (10, 506, 507). 

Lead inhibits PBG synthase and ferrochelatase, restricting haem biosynthesis and resulting in microcytic hypochromic anaemia and porphyria. Urinary excretion of 5-ALA is increased. 

Methionine deficiency leads to coproporphyrin accumulation. Lascelles and Hatch [145] suggested that heme formation may be inhibited under these conditions, perhaps at the iron insertion step because methionine is required for the synthesis of phosphatidyl choline and the latter appears to be needed for ferrochelatase activity. Tait [147a] reported that under anaerobic conditions the conversion of coproporphyrinogen to protoporphyrinogen required methionine, ATP, and ferrous ions. 

Two enzyme systems have shown themselves to be extremely sensitive to lead at low levels. The first of these is D-amino laevulinic acid dehydratase (d-ALAD), the initial and rate limiting step in the porphyrin synthetic pathway. In both experimental (Barlow et al., 1977) and clinical studies (Piomelli et al., 1980) this enzyme is potently inhibited by lead. In a review of dose-dependent low level lead effects, Zielhuis (1975) calculated a no effect level for this enzyme at about 10/tg Pb/lOOml blood in man. Other enzymes in the porphyrin synthetic pathway are also affected by lead, e.g. ferrochelatase, the enzyme responsible for the insertion of haem into the porphyrin precursor protoporphyrin IX (Moore, 1975), but this is probably due to D-ALAD-related interactions. Silbergeld and Lamon (1980) have speculated that the neurotoxic effects of lead may be due in part to a competitive interaction involving amino laevulinic acid at neuronal receptors, as there may be similarities between features of lead toxicity and some of the porphyrinopathic diseases (Moore et al., 1980). 

Another enzyme, ferrochelatase, is also inhibited at low blood lead levels. Inhibition of ferrochelatase leads to increased free erythrocyte protoporphyrin (FEP) in the blood which can then bind to zinc to yield zinc protoporphyrin. At a blood lead level of 50 [ig/dl or greater, nearly 100 percent of the population will have an increase in FEP. There is also an exponential relationship between blood lead levels greater than 40 [ig/dl and the associated ZPP level, which has led to the development of the ZPP screening test for lead exposure. 

This section deals with the long-recognized effects of Pb exposure on heme biosynthesis, specifically with reference to accumulation of heme intermediates in various organs, tissues, and biomarker media. The topic has been extensively reviewed in individual and consensus tracts (NAS/NRC, 1972, 1993 U.S. ATSDR, 2007 U.S. CDC, 1978, 1985, 1991, 2005 U.S. EPA, 1977, 1986, 2006 Waldron 1966). Lead impairs heme biosynthesis at various steps with a particularly significant effect in accumulation of protoporphyrin IX (EP, FEP, ZPP), analyzed as either the metal-free (FEP) or zinc (ZPP) complex. Other measured effects in the biosynthetic path include potent inhibition of the enzyme 6-ALA-D, moderate inhibition of the enzyme coproporphyrinogen oxidase, and relatively potent inhibition of the enzyme ferrochelatase. Associated occurrences following inhibitory effects on these enzymes include accumulation of 8-ALA in plasma (6-ALA-P), various organs and tissues, and urine (6-ALA-U), accumulation of coproporphyrin in urine (CP-U), and stimulation of the enzyme 6-ALA synthetase (6-ALA-S) in derepressive, feedback response to accumulation of 6-ALA and EP. 

An alternate mechanism to explain the etiology of elevated erythrocyte ZPP during lead exposure has its basis in the relative affinities of iron and zinc as substrates for ferrochelatase. Ferrous iron is the preferential substrate of ferrochelatase and is also an effective inhibitor of zinc utilization by this enzyme. However, when the concentration of iron as Fe decreases to suboptimal levels, as in iron deficiency, zinc is utilized by ferrochelatase as a substrate. Studies have suggested that lead decreases the availability of Fe as a substrate for ferrochelatase by inhibiting the enzymatic reduction of Fe " to Fe within mitochondria (Taketani et al. 1985), as required for use... 

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